Alignment-Free k-mer Analysis Reveals Age-Associated k-mer Signals in Canine Buccal DNA and Cell-Free DNA
- 1University of California, Los Angeles
Abstract
Aging is associated with progressive molecular and cellular changes that alter chromatin organization, epigenetic regulation, and microbial community structure. While DNA methylation-based biomarkers have demonstrated age-prediction performance, the potential of alignment-free sequence analysis for detecting age-associated signals remains unexplored. We performed a k-mer-based analysis of whole-genome sequencing datasets from two canine cohorts: 1,187 buccal DNA samples and 100 cell-free DNA (cfDNA) samples. Age-associated k-mers were identified through correlation analysis, and sequence motifs and nucleotide composition patterns were characterized across both sample types. Ridge regression models were used to predict age from k-mer frequency profiles, and synthetic datasets were generated under biological compositional assumptions to evaluate potential drivers of the observed signals. Age-associated k-mer signals were identified in both buccal DNA and cfDNA, with substantially stronger patterns observed in buccal samples. Buccal-derived k-mers exhibited more pronounced motif patterns, stronger GC and AT enrichment trends, and improved age-prediction performance relative to cfDNA. Nucleotide enrichment trends differed between sample types, suggesting that distinct biological processes may contribute to age-associated sequence composition changes in buccal DNA and cfDNA. Simulation analyses successfully reproduced major enrichment trends observed empirically, and age-associated signals were largely preserved following restriction to read-start sequence composition. These findings demonstrate that alignment-free k-mer analysis can detect age-associated sequence composition signals in canine DNA and highlight its potential as a computationally efficient complementary approach for aging biomarker discovery.
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BibTeX
@misc{hu2026alignmentfree,
title = {{Alignment-Free k-mer Analysis Reveals Age-Associated k-mer Signals in Canine Buccal DNA and Cell-Free DNA}},
author = {S. Hu and G. D. Vecchio and M. Dlugosz and M. Kokot and S. Deorowicz and M. Pellegrini},
howpublished = {bioRxiv},
year = {2026},
month = oct,
doi = {10.64898/2026.10.24.761006},
url = {https://www.biorxiv.org/content/10.64898/2026.10.24.761006v1},
note = {CC BY},
}
RIS
TY - UNPB
TI - Alignment-Free k-mer Analysis Reveals Age-Associated k-mer Signals in Canine Buccal DNA and Cell-Free DNA
AU - S. Hu
AU - G. D. Vecchio
AU - M. Dlugosz
AU - M. Kokot
AU - S. Deorowicz
AU - M. Pellegrini
PY - 2026
DA - 2026/10/24
PB - bioRxiv
DO - 10.64898/2026.10.24.761006
UR - https://www.biorxiv.org/content/10.64898/2026.10.24.761006v1
AB - Aging is associated with progressive molecular and cellular changes that alter chromatin organization, epigenetic regulation, and microbial community structure. While DNA methylation-based biomarkers have demonstrated age-prediction performance, the potential of alignment-free sequence analysis for detecting age-associated signals remains unexplored. We performed a k-mer-based analysis of whole-genome sequencing datasets from two canine cohorts: 1,187 buccal DNA samples and 100 cell-free DNA (cfDNA) samples. Age-associated k-mers were identified through correlation analysis, and sequence motifs and nucleotide composition patterns were characterized across both sample types. Ridge regression models were used to predict age from k-mer frequency profiles, and synthetic datasets were generated under biological compositional assumptions to evaluate potential drivers of the observed signals. Age-associated k-mer signals were identified in both buccal DNA and cfDNA, with substantially stronger patterns observed in buccal samples. Buccal-derived k-mers exhibited more pronounced motif patterns, stronger GC and AT enrichment trends, and improved age-prediction performance relative to cfDNA. Nucleotide enrichment trends differed between sample types, suggesting that distinct biological processes may contribute to age-associated sequence composition changes in buccal DNA and cfDNA. Simulation analyses successfully reproduced major enrichment trends observed empirically, and age-associated signals were largely preserved following restriction to read-start sequence composition. These findings demonstrate that alignment-free k-mer analysis can detect age-associated sequence composition signals in canine DNA and highlight its potential as a computationally efficient complementary approach for aging biomarker discovery.
ER -